Short interfering nucleic acid (siNA) compositions

Abstract

The present invention relates to compounds, compositions, and methods for the study, diagnosis, and treatment of traits, diseases and conditions that respond to the modulation of gene expression and/or activity, and/or modulate a gene expression pathway. Specifically, the invention relates to double-stranded nucleic acid molecules including small nucleic acid molecules, such as short interfering nucleic acid (siNA) molecules that are capable of mediating or that mediate RNA interference (RNAi) against target gene expression.

The invention claimed is: 
     
       1. A composition comprising:
 a double-stranded short interfering nucleic acid (siNA) molecule that modulates the expression and/or activity of a target RNA sequence, wherein the molecule has a passenger strand and a guide strand, wherein the guide strand comprises the structure represented by the formula: 
 
       
         
           
                 
               
                   3′-  N   S   N   N  N  N  N  N  N  N  N  N  N  N  N  N  N  N  N  S   N   S N  S   N   
                 
             
                
               
            
           
         
         wherein, 
         (a) the guide strand is complementary to the target sequence, wherein one or more mismatches between the guide strand and the target sequence are tolerated so long as its activity is maintained; 
         (b) every N nucleotide is a 2′-deoxy-2′-fluoro nucleotide and every N nucleotide is a 2′-O-methyl nucleotide; and 
         (c) each S is a phosphorothioate or phosphorodithioate internucleotide linkage. 
       
     
     
       2. The composition according to  claim 1 , wherein the passenger strand comprises the structure represented by the formula: 
       
         
           
                 
               
                   5′- B 1 -N  N  N  N  N  N  N  N  N  N  N  N  N  N  N  N  N  N  N  N   S   N - 
                 
                     
                 
                   B 2   
                 
             
                
                
                
               
            
           
         
         wherein, 
         (a) B 1  and B 2 , are each independently a terminal cap optionally including a ligand, polymer, protein or peptide transduction domain, nuclear localization sequence, cell penetrating peptide, receptor, steroid, vitamin, antibody, protamine, and/or hormone, optionally attached via a linker; 
         (b) every N nucleotide in the passenger strand is a 2′-deoxy-2′-fluoro nucleotide and every N nucleotide in the passenger strand is a 2′-O-methyl nucleotide; and 
         (c) S in the passenger strand is a phosphorothioate or phosphorodithioate internucleotide linkage. 
       
     
     
       3. The composition according to  claim 1 , wherein the double stranded siNA comprises the structure represented by formula (K): 
       
         
           
                 
               
                   5′- B 1 -N  N  N  N  N  N  N  N  N  N  N  N  N  N  N  N  N  N  N  N   S   N - 
                 
                     
                 
                   B 2   
                 
                     
                 
                   3′-  N   S   N N  N  N  N  N  N  N  N  N  N  N  N  N  N  N  N  S   N   S N  S   N   
                 
                     
                 
                   (K) 
                 
             
                
                
                
                
                
                
                
               
            
           
         
         wherein the upper strand is the passenger strand and the lower strand is the guide strand of the double-stranded nucleic acid molecule; the passenger strand is complementary to the guide strand, wherein one or more mismatches between the guide strand and passenger strand are tolerated so long as RNAi activity is maintained. 
       
     
     
       4. The composition according to  claim 2 , wherein B 1  comprises: 
       
         
           
           
               
               
           
         
         wherein each L is independently a linker and GalNAc is an N-acetyl galactosamine moiety. 
       
     
     
       5. The composition according to  claim 2 , wherein B 1  comprises: 
       
         
           
           
               
               
           
         
       
     
     
       6. The composition according to  claim 2 , wherein B 1  comprises: 
       
         
           
           
               
               
           
         
         wherein n is an integer from 1 to 20. 
       
     
     
       7. The composition according to  claim 2  wherein B 1  or B 2  comprises: 
       
         
           
           
               
               
           
         
         wherein X is O—, —S—, —CH 2 — or —NH—; n is 1, 2, 3, or 4; and the bond with “.” indicates the point of attachment optionally including one or more linkers, which may be the same or different; and further optionally including one or more targeting ligands, solubilizing agents, pharmacokinetics enhancing agents, lipids, and/or masking agents. 
       
     
     
       8. The composition according to  claim 1 , further comprising a terminal phosphate group at the 5′-end of the guide strand. 
     
     
       9. The composition according to  claim 8 , wherein the terminal phosphate group comprises a phosphate group, a diphosphate group, or a triphosphate group. 
     
     
       10. The composition according to  claim 1 , wherein 5, 6, 7, 8, or more of the 2′-O-methyl N nucleotides are pyrimidines. 
     
     
       11. The composition according to  claim 1 , wherein 5, 6, 7, 8, or more of the 2′-deoxy-2′-fluoro N nucleotides are purines. 
     
     
       12. The composition according to  claim 1 , wherein each S is a phosphorothioate internucleotide linkage. 
     
     
       13. The composition according to  claim 1 , wherein each S is a phosphorodithioate internucleotide linkage. 
     
     
       14. The composition according to  claim 1 , wherein one or more of the N nucleotides comprises a ribose sugar moiety. 
     
     
       15. The composition according to  claim 1 , wherein one or more of the N nucleotides comprises a modified sugar moiety. 
     
     
       16. The composition according to  claim 15 , wherein in the modified sugar moiety is an acyclic ribose sugar. 
     
     
       17. The composition according to  claim 1 , wherein one or more of the N nucleotides comprises a ribose sugar moiety. 
     
     
       18. The composition according to  claim 1 , wherein one or more of the N nucleotides comprises a modified sugar moiety. 
     
     
       19. The composition according to  claim 18 , wherein in the modified sugar moiety is an acyclic ribose sugar. 
     
     
       20. The composition according to  claim 1 , wherein the target RNA sequence is a viral RNA or an endogenous RNA of a human cell. 
     
     
       21. A composition comprising the double-stranded short interfering nucleic acid (siNA) molecule according to  claim 1 , and a pharmaceutically acceptable carrier or diluent. 
     
     
       22. The composition according to  claim 2 , wherein B 1  or B 2  comprise an inverted abasic 2-deoxyribose moiety. 
     
     
       23. The composition according to  claim 2 , wherein B 1  and B 2  each comprise an inverted abasic 2-deoxyribose moiety. 
     
     
       24. The composition according to  claim 2 , wherein B 1  comprises one or more galactosamine moieties optionally attached via a linker to an inverted abasic 2 deoxyribose moiety, and B 2  comprises an inverted abasic 2-deoxyribose moiety. 
     
     
       25. The composition according to  claim 2 , wherein B 1 comprises a ligand, polymer, protein or peptide transduction domain, nuclear localization sequence, cell penetrating peptide, receptor, steroid, vitamin, antibody, protamine, and/or hormone, optionally attached via a linker to an inverted abasic 2-deoxyribose moiety; and B 2  comprises a ligand, polymer, protein or peptide transduction domain, nuclear localization sequence, cell penetrating peptide, receptor, steroid, vitamin, antibody, protamine, and/or hormone, optionally attached via a linker to an inverted abasic 2-deoxyribose moiety.